The project will examine proliferation, metabolism, and interactions of retinal vascular cells by means of an established tissue culture model. Knowledge of the contributions of each cell type to the functional and structural integrity of the retinal circulation is important for understanding and treating retinal neovascularization in general and diabetic retinopathy in particular. These studies may lead to the discovery of agents capable of regulating retinal vascullr cells in vivo. Optimal tissue culture conditions will be established to allow growth of cloned lines of retinal vascular endothelia cells, pericytes, and smooth muscle cells from bovine, normal human, and diabetic human eyes. Cultured retinal vascular cells will be examined for the presence of two identifying markers of endothelia cells, factor VIII antigen and angiotensin-converting enzyme, using indirect immunofluorescence and a radioassay, respectively. Growth response of retinal capillary and retinal large-vessel cells to agents such as fibroblast growth factor, endothelial cell growth supplement, tumor-conditioned medium, vitreous extracts, and retinal growth factor will be analyzed from total cell counts. This study will also characterize the proliferative capacity of retinal vascular cells at various cell densities. The production of sorbitol and basement membrane material by cultured retinal vascular cells is pertinent to the study of diabetes. Synthesis of sorbitol by retinal vascular endothelial cells, effect of high-glucose medium and aldose reductase inhibitors upon sorbitol accumulation, cell growth kinetics, cell ultrastructure, and basement membrane production will be assessed. The extracellular material will be analyzed with radiaoactive collagen and glycoprotein precursors of basement membrane matrix. Possible interations among retinal vascular cells will be examined by demonstration of fibronectin within basement membrane matrix using indirect immunofluorescence and by construction of an "artificial retina-vessel wall" in tissue culture. The latter will allow evaluation of the "cell feeder" function of retinal pericytes and smooth muscle upon endothelial cell proliferation and synthesis of basement membrane materials. These studies are expected to help elucidate the possible role of retinal vascular cells in the pathogenesis of diabetic retinopathy.